Using neuroblastoma as a model system, we propose to establish that two categories of chromosomal instability (sites susceptible to breakage and rearrangement in the karyotype) play a significant role in the development of human cancer. The first category includes deletions/rearrangements resulting in the loss of structural material: these are postulated to point to the locations of genes responsible for tumorigenesis. The second category includes homogenously staining regions (HSRs) and double minutes (DMS), both sites of gene amplification: it is hypothesized that these contribute to tumor progression. Recombinant DNA probes of two types-those that recognize restriction fragment length polymorphisms (RFLPs) along specific chromosomes and those containing sequences amplified in HSR/DMS- will be used (a) to determine whether homozygosity for gene loci on a particular chromosome is involved in neuroblastoma tumorigenesis, and (b) to gain insight into the mechanisms that underlie gene amplification and the formation of HSR/DMS. Several of the probes to be used have been made available to us; others will be generated from genomic DNA libraries that we are constructing. If (a) is shown to be correct, we will have mapped a gene responsible for tumorigenesis in man to a specific chromosome (the third such assignment, after the genes for retinoblastoma and Wilms' tumor). In the case of (b), we will determine whether sub-microscopic rearrangements and transposition of DNA sequences can precede gene amplification. If so, it is postulated that the site of origin of the sequences participating in HSR formation in neuroblastoma (and, presumably, in other tumors as well) can be incuded in a new category of "fragile sites." The latter are hypothesized to constitute those sequences which, as the result of recombination to produce a transcriptionally active gene, induce instability in the chromosome and predispose to breakage and translocation within the karyotype.